1. Field of the Invention
The present invention relates generally to the field of genes and proteins derived from pathogenic bacteria. More particularly, the invention provides Acinetobacter outer membrane protein and gene compositions and methods for making and using a range of biological components related thereto. The invention thus provides nucleic acids, proteins, peptides and antibodies for use in various diagnostic and therapeutic applications, including those connected with peptic ulcers and cancers, respiratory diseases, sepsis and other conditions.
2. Description of Related Art
Helicobacter pylori (H. pylori) causes chronic gastritis and is correlated with the development of peptic ulcer disease and gastric carcinoma (Marshall et al, 1994; Graham, 1998; Nomura et al., 1991). However, there is also a potential association between colonization of the stomach by non-Helicobacter organisms and gastric atrophy and gastritis (Elliott et al., 1998; Haruma et al., 1995; Saunders et al., 1998).
Studies in the human and monkey have clearly shown that bacteria are important in triggering mucosal damage and inflammation in the stomach (Khanolkar-Gaitonde et al., 2000; Stockbruegger et al., 1984). It is not currently known whether colonization by non-Helicobacter organisms triggers perturbations in the neuroendocrine and epithelial cell populations. The implications being that the pathology observed may not be specific for H. pylori, but instead is the general response of the gastric mucosa to bacteria.
H. pylori is characterized by its ability to survive in the low-pH environment of the stomach by generating an alkaline microenvironment. However, with reduced levels of acid (hypochlorhydria or achlorhydria), the competitive niche established by H. pylori dissipates and the human stomach is colonized by other organisms (Haruma et al., 1995; Lehy et al., 2000). The colonization with aerobic and anaerobic flora that occurs in the stomach with increasing pH (Torres et al., 1996) may be the result of increasing age, malnutrition or iatrogenically induced achlorhydria, e.g., H2-receptor blockade or proton-pump inhibitor administration. Importantly, chronic achlorhydria is a risk factor for gastric cancer (Seery, 1991). With the exception of xe2x80x9cstress ulcerxe2x80x9d treatment, the bacterial flora present under conditions of hypochlorhydria has been poorly studied.
Patients in intensive care units often have a gastric pH of  greater than 3 due to the routine use of antacids, proton pump inhibitors or H2-receptor antagonists to prevent xe2x80x9cstress ulcersxe2x80x9d. An increase in gastric pH permits colonization of the stomach with opportunistic pathogens that contribute to the development of nosocomial pneumonia (Garrouste-Orgeas et al., 1997). Bacteria cultured from the relatively alkaline stomachs of ventilated patients (Garrouste-Orgeas et al, 1997) have been implicated in nosocomial respiratory infections (Craven et al., 1990). In a study examining oropharyngeal and gastric colonization using DNA genomic analysis, gastric colonization occurred regardless of the pH, which ranged from 2.8 to 5.7. Antacids were not used and H2-receptor antagonists were used occasionally. A notable incidence if nosocomial pneumonias was reported in ventilated patients, despite the use of broad-spectrum antibiotics, e.g., amoxicillin and aminoglycosides.
As pathogenic organisms evidently exist that are able to evade or counteract currently available antibiotics, there is a need in the art to identify organisms and virulence factors from gastric bacterial flora that cause or contribute to gastric and systemic diseases. The identification of a surface accessible molecule from such a pathogenic organism would be a significant advance, leading long sought after diagnostics and therapeutics.
The present invention overcomes these and other drawbacks inherent in the prior art by linking Acinetobacter with various diseases and disorders and by providing Acinetobacter outer membrane protein A (OMP A) protein and gene compositions for therapeutics and diagnostics. The OMP A nucleic acid, protein, peptide and antibody compositions thus provided are useful in a variety of embodiments, including the diagnosis and therapy of peptic ulcers and cancers, respiratory diseases, sepsis and other conditions. Uses of the invention as part of a battery of diagnostic agents and in combination therapies are also provided.
The invention thus provides isolated nucleic acid molecules or segments comprising at least a first isolated coding region of at least about 800, 850, 900, 910, 920, 930, 940, 950, 960, 970 or 980 nucleotides or so in length that specifically hybridizes, preferably under conditions of stringency, and more preferably under conditions of high stringency, to the coding region of the nucleotide sequence set forth in SEQ ID NO:1 or to the coding region of the nucleotide sequence set forth in SEQ ID NO:6.
The at least a first isolated coding regions are those wherein:
(a) the coding sequence is a coding sequence of a DNA molecule present in a bacterial gene library, wherein the DNA molecule hybridizes with a probe having the sequence of the complement of SEQ ID NO:1 under conditions of high stringency; or
(b) the coding sequence has a nucleotide sequence degenerate with a sequence according to (a), above.
The invention also provides isolated nucleic acid molecules or segments comprising at least a first isolated coding region that encodes a protein or polypeptide comprising an amino acid sequence that is at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identical to amino acids 22-349 of the amino acid sequence of SEQ ID NO:2. The encoded proteins and polypeptides are Acinetobacter outer membrane proteins and polypeptides or are able to generate antibodies that cross-react therewith.
Co-owned U.S. Pat. No. 6,074,840 is specifically incorporated herein by reference for purposes of supplementing the present description and enabling teaching concerning isolated nucleic acid molecules that comprise sequences encoding contiguous amino acids sequences; that encode proteins or polypeptides that exhibit at least 90% identity to a given amino acid sequence, wherein the proteins or polypeptides maintain the function of the protein or polypeptide of the given amino acid sequence; and that encode nucleic acid molecules that comprise the nucleotide sequence of a coding sequence of a DNA molecule present in a gene library, wherein the DNA molecule hybridizes with a probe having a given sequence under conditions of high stringency or nucleotide sequences degenerate with such hybridizing sequences.
Exemplary nucleic acid segments of the invention comprise at least a first isolated coding region of at least about 980 nucleotides in length that specifically hybridizes to the nucleotide sequence between nucleotide 1 and about 1050 of SEQ ID NO:1 or an isolated coding region that encodes a protein that comprises at least amino acids 22-349 of the amino acid sequence of SEQ ID NO:2.
Further nucleic acid segments comprise at least a first isolated coding region of at least about 980 nucleotides in length that specifically hybridizes to the nucleotide sequence between nucleotide 63 and about 1050 of SEQ ID NO:1 or an isolated coding region that encodes a protein that comprises at least amino acids 22-349 of the amino acid sequence of SEQ ID NO:2.
Other nucleic acid segments comprise at least a first isolated coding region of at least about 1050 nucleotides in length that specifically hybridizes to the coding region within the nucleotide sequence of SEQ ID NO:1 or to the coding region of the nucleotide sequence set forth in SEQ ID NO:6 or an isolated coding region that encodes a protein comprising an amino acid sequence that is at least about 90% identical to the amino acid sequence of SEQ ID NO:2.
Still further nucleic acid segments comprise at least a first isolated coding region of at least about 1050 nucleotides in length that specifically hybridizes to the nucleotide sequence between nucleotide 1 and about 1050 of SEQ ID NO:1 or an isolated coding region that encodes a protein having the amino acid sequence of SEQ ID NO:2.
Yet other nucleic acid segments comprise at least a first isolated coding region that encodes an Acinetobacter outer membrane protein, wherein the isolated coding region specifically hybridizes to the coding region set forth in the nucleotide sequence of SEQ ID NO:1 or encodes a protein having an amino acid sequence that is at least about 90% identical to the amino acid sequence of SEQ ID NO:2.
The nucleic acid segments also comprise at least a first isolated coding region that encodes an Acinetobacter outer membrane protein, wherein the isolated coding region specifically hybridizes to the nucleotide sequence between nucleotide 1 and about 1050 of the nucleotide sequence of SEQ ID NO:1 or to the coding region of the nucleotide sequence set forth in SEQ ID NO:6 or that encodes a protein having an amino acid sequence that is at least about 95% identical to the amino acid sequence of SEQ ID NO:2.
Certain nucleic acid segments comprise at least a first isolated coding region that encodes an Acinetobacter outer membrane protein, wherein the isolated coding region specifically hybridizes to the nucleotide sequence before the stop codon of SEQ ID NO:1 or encodes a protein comprising the amino acid sequence of SEQ ID NO:2.
Other nucleic acid segments comprise at least a first isolated coding region that encodes an Acinetobacter outer membrane protein A (OMP A), wherein the isolated coding region specifically hybridizes to the nucleotide coding sequence within SEQ ID NO:1 under stringent hybridization conditions or encodes a protein having the amino acid sequence of SEQ ID NO:2.
Certain preferred first isolated coding regions specifically hybridize to the nucleotide sequence of SEQ ID NO:1 under stringent hybridization conditions. Other preferred first isolated coding regions encode a polypeptide or protein having the amino acid sequence of SEQ ID NO:2. The isolated coding regions may have the nucleotide sequence of SEQ ID NO:1 or SEQ ID NO:6. The nucleic acid segments may be RNA or DNA segments.
The nucleic acid segments may further comprise at least a second isolated coding region that encodes a second peptide or protein, such as a second Acinetobacter peptide or protein. Such first isolated coding regions are operatively attached, in frame, to a second coding region that encodes a selected peptide or protein sequence, the nucleic acid segment encoding a fusion protein in which the Acinetobacter outer membrane protein is linked to the selected peptide or protein.
Acinetobacter bacteria that lack a functional endogenous nucleic acid that encodes an outer membrane protein having the amino acid sequence of SEQ ID NO:2 are also provided. Such bacteria include gene knockouts and other forms of genetic inactivation.
The first isolated coding regions are preferably positioned under the control of a promoter. This includes wherein the first isolated coding region is positioned, in reverse orientation, under the control of a promoter that directs the expression of an antisense product.
Thus, the present invention further provides recombinant vectors and recombinant host cells. Exemplary expression vectors comprise at least a first expression unit that comprises a promoter that expresses at least a first isolated coding region that encodes an Acinetobacter outer membrane protein, wherein the isolated coding region specifically hybridizes to the nucleotide sequence of SEQ ID NO:1 or encodes a protein comprising an amino acid sequence that is at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identical to amino acids 22-349 of the amino acid sequence of SEQ ID NO:2.
The recombinant host cells comprising at least a first exogenous nucleic acid segment that comprises at least a first isolated coding region that encodes an Acinetobacter outer membrane protein, wherein the isolated coding region specifically hybridizes to the nucleotide sequence of SEQ ID NO:1 or encodes a protein comprising an amino acid sequence that is at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identical to amino acids 22-349 of SEQ ID NO:2. Prokaryotic, eukaryotic and mammalian host cells are included.
The exogenous nucleic acid segment is preferably introduced into the cell by means of a recombinant vector and the host cell preferably expresses the exogenous nucleic acid segment to produce the encoded Acinetobacter outer membrane protein. In certain cases, the at least a first isolated coding region encodes an Acinetobacter outer membrane protein that further comprises sufficient amino acids from amino acids 1-22 of SEQ ID NO:2 to function as a signal peptide.
Method of using a nucleic acid segment that encodes an Acinetobacter outer membrane protein are provided, comprising expressing the nucleic acid segment in a recombinant host cell; wherein the nucleic acid segment comprises at least a first isolated coding region that specifically hybridizes to the nucleotide sequence of SEQ ID NO:1 or that encodes a protein comprising an amino acid sequence that is at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identical to amino acids 22-349 of SEQ ID NO:2. Met producing an Acinetobacter outer membrane proteins comprise similar process steps.
The methods may further comprise collecting the expressed Acinetobacter outer membrane protein from the host cell. The isolated coding region may encode a protein that is at least about 90% identical to amino acids 1-349 of SEQ ID NO:2 and that is expressed in the outer membrane of the host cell.
Compositions comprising isolated Acinetobacter outer membrane proteins are also provided and comprise an amino acid sequence that is at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identical to amino acids 22-349 of the amino acid sequence of SEQ ID NO:2. Proteins comprising the amino acid sequence from amino acid 22 to amino acid 349 of SEQ ID NO:2 or the amino acid sequence of SEQ ID NO:2 are preferred. All such proteins may comprise at least a first labeled amino acid or is linked to a detectable labels.
Antigenic peptides of Acinetobacter outer membrane proteins are provided, as exemplified by those comprising the amino acid sequence of GSRTVLAEQPVAQ (amino acids 337-349 of SEQ ID NO:2), DTQHNNNGND (amino acids 36-45 of SEQ ID NO:2), TGNYDSKVKP (amino acids 111-120 of SEQ ID NO:2) or YKYEFEGVPRGTRGNEEEG (amino acids 129-147 of SEQ ID NO:2).
Purified antisera and antibodies are preferred aspects of the present invention. These compositions and components have immunospecificity for an Acinetobacter outer membrane protein that has an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2. Polyclonal and monoclonal antibodies are provided, as are and antibodies linked to detectable labels.
First and second nucleic acid primers are also encompassed by the invention, e.g., wherein the primers hybridize to spatially distant sequences from the nucleic acid sequence of SEQ ID NO:1 and are capable of specifically amplifying an Acinetobacter outer membrane protein nucleic acid sequence when used in conjunction with a polymerase chain reaction.
Diagnostic kits of the invention generally comprise a detection reagent in operative association with:
(a) at least a first Acinetobacter outer membrane protein-encoding nucleic acid sequence that specifically hybridizes to the nucleotide sequence of SEQ ID NO:1;
(b) a pair of nucleic acid primers that hybridize to spatially distant sequences from the nucleic acid sequence of SEQ ID NO:1, the primers being capable of specifically amplifying an Acinetobacter outer membrane protein nucleic acid sequence when used in conjunction with a polymerase chain reaction;
(c) a plurality of oligonucleotides that each have a sequence from SEQ ID NO:1, the plurality of oligonucleotides being attached to a solid support and available for hybridization;
(d) at least a first antibody that has immunospecificity for an Acinetobacter outer membrane protein having an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2; or
(e) an isolated Acinetobacter outer membrane protein comprising an amino acid. sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2.
Nucleic acid detection kits comprise a detection reagent in operative association with at least a first Acinetobacter outer membrane protein-encoding nucleic acid sequence that specifically hybridizes to the nucleotide sequence of SEQ ID NO:1.
The at least a first nucleic acid sequence may be substantially the same length as the coding region within the nucleotide sequence of SEQ ID NO:1. Primers that hybridize to spatially distant sequences from the nucleic acid sequence of SEQ ID NO:1 and that specifically amplify an Acinetobacter outer membrane protein nucleic acid sequence when used in conjunction with a polymerase chain reaction may also be used.
Multiple copies of the at least a first nucleic acid sequence may also be attached to a solid support and available for hybridization. A plurality of oligonucleotides that each have a sequence from SEQ ID NO:1 may also be attached to a solid support and available for hybridization.
As such, the present invention provides sequence detection chips, comprising at least a first Acinetobacter outer membrane protein-encoding nucleic acid sequence from SEQ ID NO:1 attached to a solid support and available for hybridization. The multiple copies of a nucleic acid molecule having the nucleotide sequence from nucleotide 1 to about 1050 of SEQ ID NO:1 are preferably arrayed on the solid support. The plurality of oligonucleotides arrayed on the solid support preferably each have a substantially distinct sequence from SEQ ID NO:1. The plurality of oligonucleotides arrayed on the solid support may also have substantially distinct sequences from SEQ ID NO:1 that overlap by one nucleotide residue per oligonucleotide, such that an overlapping array of SEQ ID NO:1-derived sequences are presented on the solid support.
Immunodetection kits of the invention generally comprise a detection reagent in operative association with an isolated Acinetobacter outer membrane protein that comprises an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2 or at least a first antibody that has immunospecificity for the isolated Acinetobacter outer membrane protein. The detection reagent may be operatively associated with or operatively attached to the first antibody; or it may be operatively attached to a second antibody that has immunospecificity for the first antibody.
Antigen compositions comprising biologically effective amounts of at least an isolated Acinetobacter outer membrane protein are provided. The proteins generally comprises an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2. Antigenic peptides derived from the Acinetobacter outer membrane protein are also included.
Antigenic cocktails comprise a combined immunogenic amount of an isolated Acinetobacter outer membrane protein that comprises an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% of 99% or so identity to amino acids 22-349 of SEQ ID NO:2, or an antigenic peptide from the Acinetobacter outer membrane protein, and at least a second antigenic component.
The at least a second antigenic component may be an Acinetobacter antigenic component or an H. pylori antigenic component, such as an H. pylori CagA antigenic component or an H. pylori VacA antigenic component. At least one adjuvant may be included.
Vaccine formulations likewise comprise, in a pharmaceutically acceptable form, a therapeutically effective amount of at least an isolated Acinetobacter outer membrane protein that comprises an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2, an antigenic peptide from the Acinetobacter outer membrane protein; or an isolated nucleic acid that encodes the Acinetobacter outer membrane protein or an antigenic peptide therefrom.
In terms of methods, the present invention provides methods for detecting a nucleic acid that encodes an Acinetobacter outer membrane protein, comprising contacting sample nucleic acids suspected of containing a nucleic acid that encodes an Acinetobacter outer membrane protein with at least a first isolated nucleic acid that specifically hybridizes to at least a first portion of the nucleotide sequence of SEQ ID NO:1, under conditions effective to allow hybridization of substantially complementary nucleic acids, and detecting the hybridized complementary nucleic acids thus formed.
Methods for detecting an Acinetobacter outer membrane protein comprise contacting a sample suspected of containing an Acinetobacter outer membrane protein with an antibody that has immunospecificity for an Acinetobacter outer membrane protein having an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2, under conditions effective to allow the formation of immune complexes, and detecting the immune complexes thus formed.
Methods for identifying the presence of Acinetobacter in a biological sample comprise testing a biological sample suspected of containing Acinetobacter for the presence of a protein with 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so sequence identity to SEQ ID NO:2, wherein the presence of the protein is indicative of a sample that contains Acinetobacter. The presence of the protein may be identified by a molecular biological assay to identify a nucleic acid that encodes the protein; by an immunoassay to identify the protein.
The methods include those for diagnosing a medical or veterinary condition associated with Acinetobacter and comprise testing a biological sample from an animal or human for the presence of a nucleic acid that specifically hybridizes to SEQ ID NO:1, for the presence of a protein with 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so sequence identity to amino acids 22-349 of SEQ ID NO:2, or for the presence of an antibody that has immunospecificity for the protein, wherein the presence of the nucleic acid, protein or antibody is indicative of a medical or veterinary condition associated with Acinetobacter.
In testing for the presence of the nucleic acid segment, the method may comprise the steps of:
(a) contacting nucleic acids of the biological sample with at least a first isolated nucleic acid that specifically hybridizes to at least a first portion of the nucleotide sequence of SEQ ID NO:1, under conditions effective to allow hybridization of substantially complementary nucleic acids; and
(b) detecting the hybridized complementary nucleic acids thus formed.
The nucleic acids may be maintained within the biological sample during contact with the at least a first isolated nucleic acid or separated from the biological sample prior to contact with the at least a first isolated nucleic acid. The nucleic acids of the biological sample may be DNA or RNA.
The at least a first isolated nucleic acid may comprise a radio, enzymatic or fluorescent detectable label and the hybridized complementary nucleic acids may be detected by detecting the label.
Equally, the method may comprise the steps of:
(a) contacting nucleic acids of the biological sample with a pair of nucleic acid primers that specifically hybridize to spatially distant sequences from SEQ ID NO:1, the primers capable of amplifying a nucleic acid segment from SEQ ID NO:1 when used in conjunction with a polymerase chain reaction;
(b) conducting a polymerase chain reaction to create amplification products; and
(c) detecting the amplification products thus formed.
Further methods comprise the steps of:
(a) applying the nucleic acids of the biological sample to a solid support that presents an array of isolated nucleic acids from SEQ ID NO:1, under conditions effective to allow hybridization of substantially complementary nucleic acids; and
(b) detecting the hybridized substantially complementary nucleic acids thus formed.
The solid support may present a repeated array of the nucleic acid sequence of SEQ ID NO:1; or a plurality of oligonucleotides that each have a substantially distinct sequence so that the plurality covers the entire coding region of SEQ ID NO:1.
Where the biological sample is tested for the presence of the protein, the method may comprise the steps of:
(a) contacting the biological sample with at least a first antibody that specifically binds to a protein having at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to SEQ ID NO:2, under conditions effective to allow the formation of immune complexes; and
(b) detecting the immune complexes so formed.
The immune complexes may be detected by a detectable label linked to the first antibody or by a detectable label linked to a second antibody that specifically binds to the first antibody.
Where the biological sample is tested for the presence of the antibody, the methods may comprise the steps of:
(a) contacting the biological sample with a labeled protein having at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to SEQ ID NO:2, under conditions effective to allow the formation of labeled immune complexes; and
(b) detecting the immune complexes so formed by immunoprecipitating the labeled immune complexes.
The biological sample may be a blood, serum, sputum, stool, biopsy or endoscopy sample; e.g., an endoscopic gastric biopsy sample or a sputum or lung biopsy sample.
The biological sample may be obtained from an animal or patient suspected of having a gastrointestinal ulcer, a peptic ulcer; gastrointestinal cancer, such as gastric lymphoma or adenocarcinoma; a more alkaline gastric pH than normal; gastric atrophy or non-H. pylori chronic gastritis; a respiratory disorder; pneumonia; meningitis; pernicious anemia or sepsis. The biological sample could be obtained from an immunocompromised patient, such as an immunocompromised patient in preparation for, during or subsequent to a bone marrow or organ transplant, or from a hospitalized patient in ICU.
Methods for diagnosing veterinary or medical conditions associated with Acinetobacter thus comprise determining the amount of an Acinetobacter OMP A component present within a biological sample from an animal or patient suspected of having an Acinetobacter-associated veterinary or medical condition, wherein an increased amount of the OMP A component, in comparison to the amount within a sample from a healthy animal or subject, is indicative of an animal or patient with a veterinary or medical condition associated with Acinetobacter; wherein the Acinetobacter OMP A component is a nucleic acid that specifically hybridizes to the nucleotide sequence of SEQ ID NO:1 or a protein comprising an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2.
Treatment methods include those for generating an immune response, comprising administering to an animal a pharmaceutically acceptable composition comprising an immunologically effective amount of an isolated Acinetobacter outer membrane protein of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2; an antigenic peptide from the Acinetobacter outer membrane protein; or an isolated nucleic acid that encodes the Acinetobacter outer membrane protein or an antigenic peptide thereof. Methods wherein the pharmaceutically acceptable composition is administered intravenously are preferred, and adjuvants may be included.
The animal or patient may have, be suspected of having, or be at risk for developing,: a gastrointestinal ulcer, a peptic ulcer; gastrointestinal cancer, such as gastric lymphoma or adenocarcinoma; a more alkaline gastric pH than normal; gastric atrophy or non-H pylori chronic gastritis; a respiratory disorder; pneumonia; meningitis; pernicious anemia or sepsis.
The animal or patient may be an immunocompromised animal or patient, such as an immunocompromised patient in preparation for, during or subsequent to a bone marrow or organ transplant, or from a hospitalized patient that is or is at risk for being in critical condition.
As many OMPs have antimicrobial and antibacterial activities, the present invention further provides methods for killing a microorganism, comprising contacting a microorganism with a toxic amount of a composition comprising an isolated Acinetobacter outer membrane protein that is at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identical to amino acids 22-349 of SEQ ID NO:2. The microorganism may be one that infects the respiratory or gastrointestinal tract.
Non-biomedical compositions and methods of using this invention also exist. In particular, the present invention provides formulations and uses of Acinetobacter OmpA as at least one of the active ingredients in a bioemulsifier or biosurfactant preparation for combating oil slicks and/or for obtaining residuals from oil wells. Although not suggested in the art, the inventor reasons that these compositions and uses are analogous to those of Pseudomonas aeruginosa. 
Now that the advantageous compositions of the invention have been provided for such use, various technical skills exist in the art for ready application to stabilizing emulsions of high viscosity hydrocarbons, including crude oil and other commercial oil preparations. For example, U.S. Pat. Nos. 6,060,287 and 5,866,376 are specifically incorporated herein by reference for purposes of supplementing the present description and enabling teaching concerning the use of bacterial compositions as biosurfactants.
Accordingly, the invention provides biosurfactant formulations, preferably storage-stable formulations, that comprise at least a first isolated Acinetobacter outer membrane protein that comprises an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2, or bioemulsifying portions, domains or peptides thereof. Recombinant proteins or variants are preferred. Apparatus and kits for the application, and preferably, widespread application of such biosurfactant, Acinetobacter outer membrane protein formulations are also provided.
These formulations may be combined with other agents to combat oil spills in the environment, particularly at sea, but also including within soil samples, aquifers and the like. U.S. Pat. Nos. 5,551,987 and 5,128,262 are also specifically incorporated herein by reference for purposes of supplementing the present description and enabling teaching concerning the use of biological agents as biosurfactants for the treatment of mud and oily residues obtained from the processing or extraction of crude oil and for the microbial decontamination of soils contaminated with hydrocarbons and mineral oils.
Methods of the invention therefore include contacting a composition comprising hydrocarbons, mineral oils, high viscosity hydrocarbons and/or high viscosity crude oil with a biologically or environmentally effective amount of a biosurfactant formulation that comprises at least a first isolated Acinetobacter outer membrane protein that comprises an amino acid sequence of at least about 85%, 86%, 87%, 88%, 89%, 90%, 91% 92% 93% 94% 95% 96% 97% 98% or 99% or so identity to amino acids 22-349 of SEQ ID NO:2, or a bioemulsifying portion, domain or peptide thereof.
The xe2x80x9cbiologically or environmentally effective amountxe2x80x9d is an amount of the biosurfactant formulation or Acinetobacter outer membrane protein composition effective to stabilize hydrocarbons, mineral oils, high viscosity hydrocarbons and/or high viscosity crude oil or emulsions thereof, and/or to disperse or remove an inappropriate sample of such unwanted hydrocarbons and oils from an unintended location(s).